What is Suface Plasmon Resonance (SPR)?
The Surface Plasmon Resonance (SPR) technique is utilized to assess, in real-time, interactions between biomolecules, and in niche surface science applications. SPR systems typically consist of a microfluidic device to deliver analytes, sensor chips that create a surface to study binding, and an optical measurement system. The combination of the sensor chip and the optical measurement system are the heart of the Surface Plasmon Resonance technique, the physics of which is described below. For further information on the chemistry related to sensor chips, please visit the sensor chip page.
Technically, the term Surface Plasmon Resonance (SPR), refers to an optical phenomenon that enables monitoring of changes in refractive index via a quantum mechanical aspect. To start, it is helpful to think in terms of Snell's law.
Snell’s law describes what happens when light is directed through a high refractive index prism (e.g. Sapphire - refractive index 1.76) to a surface of the prism in contact with a low refractive index medium, for example physiological buffer (See Figure 1). Light rays below the critical angle will exit the prism bending toward the prism surface. Light rays above the critical angle are totally internally reflected back through the prism. Photons that are totally internally reflected create an electric field at the interface. Light is not coming out of the prism but an electric field extends past the reflecting surface. This field oscillates with the usual characteristics of an electromagnetic mode. The electrical component perpendicular to the interface decays exponentially – this is called an evanescent wave. This wave is bound to the surface and is used in SPR to detect changes occurring at the surface.
SPR spectroscopy adds a thin metal layer between the prism surface and the aqueous compartment (See Figure 2). Free electrons in the metal layer can act as a resonator. Energy for the resonance comes from the evanescent wave produced by the totally internally reflected photons.
When certain conditions, determined by the wavelength, illumination angle, and refractive index of the prism, metal and aqueous layers are met, coupling/resonance occurs between the plasma oscillations of the free electrons in the metal and the bound electromagnetic field of the totally internally reflected photons. This coupling is the result of the momentum of the incoming light equaling the momentum of the plasma electromagnetic field. Photons are “absorbed” and converted to surface plasmons. Since the photons are not reflected, a “shadow” occurs in the reflected light.
Reichert Inc.’s Surface Plasmon Resonance instruments are optically configured to illuminate a spot on the gold surface over a range of angles from 58 to 85 degrees. When the aqueous compartment is e.g. physiological saline solution the “shadow” or SPR minimum occurs at about 66 Degrees. Mass changes at the interface between the gold layer and the aqueous compartment cause changes in the local refractive index near the gold layer. This changes the coupling/resonance angle. For example, if a protein layer was added to the gold / aqueous interface, the coupling angle would be about 66.6 Degrees. Actual reflectivity from the SR7000 (Angle vs. reflectivity) before and after immobilizing a protein to the surface is shown in Figure 3.
In all of the spectrometers, the gold surface is illuminated with a range of angles that encompass these shifts in the coupling angle. This range of angles is continuously monitored with a linear photodiode array detector. Image analysis determines the angle at which the reflectivity minimum occurs. This is continuously sent to the data acquisition computer/software.